Aged skin commonly is afflicted by inflammatory skin diseases or xerosis/eczema that can be traced to, or exacerbated by, impaired epidermal permeability barrier homeostasis. In contrast to extremely aged individuals, we find that moderately aged humans (50-80 years) and mice (12-15 mos) suffer from defective stratum corneum (SC) acidification. SC acidification is required to activate key pH-sensitive lipid processing enzymes, including beta-GlcCer'ase (beta-glucocerebrosidase). Neonatal and adult SC both are acidified largely through the combined activity of two endogenous mechanisms, the Na+/H+ antiporter (NHE1) and the secretory phospholipase (sPLA2) enzymes, although the relative contribution of each agent in supplying SC acidity to specific regions of the SC is not known. We previously found that NHE1 protein levels and full- thickness SC acidity are decreased in moderately aged mouse epidermis. We now propose to define the relative roles of NHE1 and sPLA2 in acidifying specific regions of the SC in young adult mouse and human epidermis, and determine what role abnormal NHE1 or sPLA2 expression, localization and/or function play in producing the abnormal acidification seen in aged SC. We will define the roles of the NHE1 and sPLA2 in both murine and human skin, and then delineate the responsible mechanisms in mouse skin and cultured human epidermal equivalents. Finally, we will explore a broad set of strategies aimed at normalizing SC acidification, either by upregulating sPLA2 activity, or by bypassing defective endogenous mechanisms with exogenous acidifying agents already approved for use in humans. HYPOTHESIS: Both the NHE1 transporter and one or more sPLA2 isoforms control SC barrier bulk acidification, or acidification of SC microdomains, where the pH-sensitive lipid processing enzymes that influence barrier homeostasis are selectively and locally activated. The NHE1 and sPLA2 complement each other in acidifying different layers and microdomains within normal young adult SC. Defects in one or both acidifying mechanisms result in suboptimal function in moderately aged epidermis, depending on the severity and localization of the age-associated pH abnormality. Enhancing SC acidity, either through increased sPLA2 activity, or by applying exogenous acidifying agents, will normalize lipid processing, thereby improving the clinical abnormalities in SC function seen in aged epidermis. The short-term goal of this project is to determine the pathophysiologic mechanisms by which SC acidity differs in aged vs. young SC. The long-term goal of this project is to restore epidermal permeability barrier homeostasis in aged skin, by optimizing the SC acidity that controls barrier homeostasis. PUBLIC HEALTH RELEVANCE: Information form these experiments will be used to optimize therapeutic protocols that restore effective epidermal barrier homeostasis in aged skin. As normal skin ages, it tends to become dry. This dryness makes even normal skin itchy, and makes skin that already is affected by common diseases, such as eczema, worse. Dry skin in moderately aged people (age 50-75 years) is caused by the loss of an epidermal barrier, made up of correctly processed lipids in the uppermost layer of the skin. Processing these lipids, in turn, depends on effectively acidifying this uppermost layer of the skin. We have found that even moderately aged skin is less acidic than skin of young adults, using a new non-invasive microscopy technique called fluorescence lifetime imaging. Further, we have found that applying acidic solutions to moderately aged skin restores skin acidity and normal lipid processing, thus improving the epidermal barrier. These experiments will study what mechanisms in the skin cause its uppermost layer to be acidified in young skin, what changes occur in aged skin that cause this acidification to be lost, and what therapies work best to restore acidity and a normal epidermal barrier to aged skin. These studies will show that it is possible to identify and treat a major cause of skin disease in the aged.